Showing papers on "Quinolone published in 2003"

Development of the quinolones.

Abstract: Since their discovery in the early 1960s, the quinolone group of antibacterials has generated considerable clinical and scientific interest. Nalidixic acid, the first quinolone to be developed, was obtained as an impurity during the manufacture of quinine. Since this time, many derivatives have been synthesized and evaluated for their antibacterial potency. Two major groups of compounds have been developed from the basic molecule: quinolones and naphthyridones. Manipulations of the basic molecule, including replacing hydrogen with fluorine at position 6, substituting a diamine residue at position 7 and adding new residues at position 1 of the quinolone ring, have led to enhanced antibacterial efficacy. In general these compounds are well tolerated. However, some of these structural changes have been found to correlate with specific adverse events: the addition of fluorine or chlorine at position 8 being associated with photo-reactivity, e.g. Bay y 3118 and sparfloxacin; and the substitution of an amine or a methyl group at position 5 having a potential role in QTc prolongation, e.g. sparfloxacin and grepafloxacin. Progressive modifications in molecular configuration have resulted in improved breadth and potency of in vitro activity and pharmacokinetics. One of the most significant developments has been the improved anti-Gram-positive activity of the newer compounds, such as moxifloxacin and garenoxacin. In the current millennium, these new agents may play an important role in the treatment of respiratory infections.

Fluoroquinolones: action and resistance

Abstract: Fluoroquinolones trap gyrase and topoisomerase IV on DNA as ternary complexes that block the movement of replication forks and transcription complexes. Studies with resistant mutants indicate that during complex formation quinolones bind to a surface alpha-helix of the GyrA and ParC proteins. Lethal action is a distinct event that is proposed to arise from release of DNA breaks from the ternary complexes. Many bacterial pathogens are exhibiting resistance due to alterations in drug permeability, drug efflux, gyrase-protecting proteins, and target topoisomerases. When selection of resistant mutants is described in terms of fluoroquinolone concentration, a threshold (mutant prevention concentration, MPC) can be defined for restricting the development of resistance. MPC varies among fluoroquinolones and pathogens; when combined with pharmacokinetics, MPC can be used to identify compounds least likely to enrich mutant subpopulations. Use of suboptimal doses and compounds erodes the efficacy of the class as a whole because resistance to one quinolone reduces susceptibility to others and/or increases the frequency at which resistance develops. When using fluoroquinolones in combination therapy, the development of resistance may be minimized by optimizing regimens for pharmacokinetic overlap.

Prevalence of plasmid-mediated quinolone resistance

Abstract: Quinolone resistance encoded by the qnr gene and mediated by plasmid pMG252 was discovered in a clinical strain of Klebsiella pneumoniae that was isolated in 1994 at the University of Alabama at Birmingham Medical Center. The gene codes for a protein that protects DNA gyrase from quinolone inhibition and that belongs to the pentapeptide repeat family of proteins. The prevalence of the gene has been investigated by using PCR with qnr-specific primers with a sample of more than 350 gram-negative strains that originated in 18 countries and 24 states in the United States and that included many strains with plasmid-mediated AmpC or extended spectrum beta-lactamase enzymes. qnr was found in isolates from the University of Alabama at Birmingham only during 6 months in 1994, despite the persistence of the gene for FOX-5 beta-lactamase, which is linked to qnr on pMG252. Isolates from other locations were negative for qnr. The prevalence of mcbG in the same sample was also examined. mcbG encodes another member of the pentapeptide repeat family and is involved in immunity to microcin B17, which, like quinolones, targets DNA gyrase. A single clinical isolate contained mcbG on a transmissible R plasmid. This plasmid and one carrying the complete microcin B17 operon slightly decreased sparfloxacin susceptibility but had a much less protective effect than pMG252. Plasmid-mediated quinolone resistance was thus rare in the sample examined.

Mechanisms of quinolone resistance

Abstract: With the increasing use of quinolones for the treatment of gram-positive bacterial infections, an understanding of the mechanisms of quinolone resistance in gram-positive bacteria is of considerable importance. This chapter summarizes the current understanding of established mechanisms of resistance to this class of antimicrobial agents in gram-positive bacteria. There are important differences between gram-positive and gram negative bacteria both in target enzyme sensitivity and in the means by which efflux resistance mechanisms operate that are of clinical and fundamental importance. Quinolones interact with both of the two type 2 topoisomerases in eubacteria, DNA gyrase and topoisomerase IV, which are essential for bacterial DNA replication. Quinolone-resistant clinical and laboratory strains of Streptococcus pneumoniae have been shown to have reduced accumulation of quinolones that is reversible with reserpine, suggesting the involvement of an efflux system(s) in quinolone resistance. Quinolone-resistant clinical isolates of viridans streptococci have been shown to have an efflux phenotype defined as lower MICs of quinolones in the presence of reserpine. DNA from such strains of S. mitis and S. oralis was able to transform S. pneumoniae to efflux phenotype in the laboratory. Overexpression of norA and genes for topoisomerases from plasmids are known, however, to have toxic effects on the cell that may limit the fitness of resistant bacteria containing them. Thus, at present quinolone resistance in gram-positive bacteria is attributable exclusively to chromosomal mutations that affect quinolone targets or quinolone permeation to these targets.

In Vitro Antibacterial Potency and Spectrum of ABT-492, a New Fluoroquinolone

Abstract: ABT-492 demonstrated potent antibacterial activity against most quinolone-susceptible pathogens. The rank order of potency was ABT-492 > trovafloxacin > levofloxacin > ciprofloxacin against quinolone-susceptible staphylococci, streptococci, and enterococci. ABT-492 had activity comparable to those of trovafloxacin, levofloxacin, and ciprofloxacin against seven species of quinolone-susceptible members of the family Enterobacteriaceae, although it was less active than the comparators against Citrobacter freundii and Serratia marcescens. The activity of ABT-492 was greater than those of the comparators against fastidious gram-negative species, including Haemophilus influenzae, Moraxella catarrhalis, Neisseria gonorrhoeae, and Legionella spp. and against Pseudomonas aeruginosa and Helicobacter pylori. ABT-492 was as active as trovafloxacin against Chlamydia trachomatis, indicating good intracellular penetration and antibacterial activity. In particular, ABT-492 was more active than trovafloxacin and levofloxacin against multidrug-resistant Streptococcus pneumoniae, including strains resistant to penicillin and macrolides, and H. influenzae, including β-lactam-resistant strains. It retained greater in vitro activity than the comparators against S. pneumoniae and H. influenzae strains resistant to other quinolones due to amino acid alterations in the quinolone resistance-determining regions of the target topoisomerases. ABT-492 was a potent inhibitor of bacterial topoisomerases, and unlike the comparators, DNA gyrase and topoisomerase IV from either Staphylococcus aureus or Escherichia coli were almost equally sensitive to ABT-492. The profile of ABT-492 suggested that it may be a useful agent for the treatment of community-acquired respiratory tract infections, as well as infections of the urinary tract, bloodstream, and skin and skin structure and nosocomial lung infections.

Fluoroquinolones: structure and target sites.

Abstract: The quinolones are a potent group of drugs that target the essential bacterial enzymes DNA gyrase and topoisomerase IV. DNA gyrase is the primary target of Gram negative organisms however, it is topoisomerase IV that is the primary target of Gram positive organisms. Within these enzymes is a highly conserved region centered round the active site where resistance mutations occur. These mutations are almost always identical, irrespective of organism. In spite of the homology of this region, amino acid sequence analysis shows that there are defined differences between the Gram groups, particularly in topoisomerase IV, and it is speculated that herein lies the origin of target preference. Since the first quinolone nalidixic acid was developed, the quinolones have undergone structural modifications, in particular the addition of a fluorine at position 6, to produce the fluoroquinolones. This has seen their potency and pharmakokinetic profile greatly increase. In vitro selection of resistance mutations has allowed the observation of how resistance is acquired and some of the modifications in newer fluoroquinolones have resulted in the shift of primary target from topoisomerase IV to gyrase with Gram positives. Curiously, purified topoisomerase IV is still more sensitive even if gyrase is the primary target. Gyrase remains the primary target for Gram negatives.

Multiple antibiotic-resistance mechanisms including a novel combination of extended-spectrum β-lactamases in a Klebsiella pneumoniae clinical strain isolated in Argentina

Abstract: Klebsiella pneumoniae M1803, isolated from a paediatric patient with chronic urinary infection, presented nine antimicrobial resistance mechanisms harboured on two conjugative megaplasmids, in addition to the chromosomally mediated SHV-1 β-lactamase. These nine antimicrobial resistance mechanisms comprised two extended-spectrum β-lactamases (ESBLs) (PER-2 and CTX-M-2), TEM-1-like, OXA-9-like, AAC(3)-IIa, AAC(6')-Ib, ANT(3")-Ia and resistance determinants to tetracycline and chloramphenicol. During fluoro- quinolone treatment, a variant derived from M1803 (named M1826) was selected, with an overall increase of MICs, in particular of cefoxitin and carbapenems. No enzymic activity against these latter drugs was found. Mutations in the region analogous to the quinolone resistance-determining region were not found. Strain M1826 was deficient in OmpK35/36 expression, which produced the decrease in the susceptibility to cefoxi- tin, carbapenems and fluoroquinolones. The blaCTX-M-2 gene was located in an unusual class 1 integron, which includes Orf513, as occurred in the recently described In35. In addition, Tn3 and Tn1331 were detected in both K. pneumoniae isolates. This is the first report of in vivo selection of an OmpK35/36 deficiency in a K. pneumoniae strain that produced a novel combination of two ESBLs (CTX-M-2 and PER-2) during fluoro- quinolone treatment in a paediatric patient with chronic urinary infection.

Mechanisms of Quinolone Action

Abstract: This chapter introduces the quinolones with a brief consideration of DNA topoisomerases and quinolone target preference. Throughout the chapter attention is given to fluoroquinolone structure. Recently discovered examples include the relaxation of supercoils associated with infection of cultured macrophages by Salmonella enterica serovar Typhimurium and with hydrogen peroxide treatment of Escherichia coli. The bacterial topoisomerases are divided into three groups: type I (topoisomerases I and III), type II (gyrase and topoisomerase IV), and specialized topoisomerases (enzymes that catalyze transposition or integration/excision of bacteriophage DNA from the bacterial chromosome). A function of topoisomerase I is the topological destabilization of transcription-mediated R loops. Another is likely to be control of global supercoiling, since a topA defect that raises supercoiling also suppresses a mukB mutation, a defect that has a global effect on chromosome condensation. Quinolone binding to these mutant gyrase-DNA complexes induces a conformational change that can be detected in the GyrB subunit by limited proteolysis. The location of the quinolone-gyrase-DNA complexes on the bacterial chromosome is likely to influence the potential damage that quinolones can cause. The bacteriostatic effects of the quinolones are now understood at a level sufficient to allow structure-function interpretations. From a clinical perspective, there is a need to identify safe compounds that rapidly kill bacteria, especially resistant mutants. However, further refinement could become an academic exercise if ways are not developed to slow the emergence of fluoroquinolone-resistant pathogens.

The history of the development and changes of quinolone antibacterial agents

Abstract: The quinolones, especially the new quinolones (the 6-fluoroquinolones), are the synthetic antibacterial agents to rival the Beta-lactam and the macrolide antibacterials for impact in clinical usage in the antibacterial therapeutic field. They have a broad antibacterial spectrum of activity against Gram-positive, Gram-negative and mycobacterial pathogens as well as anaerobes. Further, they show good-to-moderate oral absorption and tissue penetration with favorable pharmacokinetics in humans resulting in high clinical efficacy in the treatment of many kinds of infections. They also exhibit excellent safety profiles as well as those of oral Beta-lactam antibiotics. The bacterial effects of quinolones inhibit the function of bacterial DNA gyrase and topoisomerase IV. The history of the development of the quinolones originated from nalidixic acid (NA), developed in 1962. In addition, the breakthrough in the drug design for the scaffold and the basic side chains have allowed improvements to be made to the first new quinolone, norfloxacin (NFLX), patented in 1978. Although currently more than 10,000 compounds have been already synthesized in the world, only two percent of them were developed and tested in clinical studies. Furthermore, out of all these compounds, only twenty have been successfully launched into the market. In this paper, the history of the development and changes of the quinolones are described from the first quinolone, NA, via, the first new quinolone (6-fluorinated quinolone) NFLX, to the latest extended-spectrum quinolone antibacterial agents against multi-drug resistant bacterial infections. NA has only modest activity against Gram-negative bacteria and low oral absorption, therefore a suitable candidate for treatment of systemic infections (UTIs) is required. Since the original discovery of NA, a series of quinolones, which are referred to as the old quinolones, have been developed leading to the first new quinolone, NFLX, with moderate improvements in over all properties starting in 1962 through and continuing throughout the 1970's. Especially, the drug design for pipemidic acid (PPA) indicated one of the important breakthroughs that lead to NFLX. The introduction of a piperazinyl group, which ia a basic moiety at the C7-position of the quinolone nuclei, improved activity against Gram-negative organisms broadening the spectrum to include Pseudomonas aeruginosa. PPA also showed soem activity against Gram-positive bac teria. The basic piperazine ring, which can form the zwitterionic natrure with the carboxylic acid at the C3-position, has subsequently been shown to increase the ability of the drugs to penetrate the bacterial cells resulting in enhanced activity. Further, the zwitterionic forms resulted in significant tissue penetration in the pharmacokinetics. On the other hand, the first compound with a fluorine atom at the C6-position of the related quinolone scaffold was flumequine and the compound indicated that activity against Gram-positive bacteria could be improved in the old quinolones. The addition of a flourine atom at the C6-position is essential for the inhibition of target enzymes. The results show the poten antibacterial activity and the penetration of the quinolone molecule into the bacterial cells and human tissue. The real breakthrough came with the combination of these two features in NFLX, a 6-fluorinated quinolone having a piperazinyl group at the C7-position, NFLX features significant differences from the old quinolones in the activities and pharmacokinetics in humans, resulting in high clinical efficacy in the treatment of many kinds of infections including RTIs.Consequently, those great discoveries are rapidly superseded by even better compounds and NFLX proved to be just the beginning of a highly successful period of research into the modifications of the new quinolone antibacterials. Simce the chemical structure and important features of NFLX had become apparent in 1978, many compounds were patented in the next three years, several of which reached the market. Among the drugs, ofloxacin (OFLX) and ciprofloxacin (CPFX) are recognized as superior in several respects to the oral beta-lactam antibiotics as an antibacterial agent. With a focus on OFLX and CPFX, numerous research groups entered the antibacterial therapeutic field, triggering intense competition in the search to find newer, more effective quinolones. After NFLX was introduced in the market, while resulting by the end of today, eleven kinds of other new quinolones launched in Japan. They are enoxacin (ENX), OFLX, CPFX, lomefloxacin (LFLX), fleroxacin (FRLX), tosufloxacin (TFLX), levofloxacin (LVFX), sparfloxacin (SPFX), gatifloxacin (GFLX), prulifloxacin (PULX) and also pazufloxacin (PZFX). The advantages of these compounds, e.g., LVFX, SPFX and GFLX, are that their spectrum includes Gram-positive bacteria species as well as Gram-negative bacteria and they improve bioavailability results when a daily dose is administered for systemic infections including RTIs. However, unexpected adverse reactions, such as the CNS reaction, the drug-drug interaction, phototoxicity, hepatotoxicity and cardiotoxicity such as the QTc interval prolongation of ECG, have been reported in the clinical evaluations or the post-marketing surveillance of several new quinolones. Moreover, the adverse reactions of arthropathy (the joint toxicity) predicated from studies in juvenile animals have never materialized in clinical use. Therefore, no drugs other than NFLX have yet been approved for pediatric use. Fortunately, the newer quinolones are being developed and tested to reduce these adverse reactions on the basis of recent studies. On the other hand, multi-drug resistant Gram-positive bacteria including methicillin-resistant Staphylococcus aureus (MRSA), methicillin-resistant coagulase-negative staphycolocci (MRCNS), penicillin-resistant Streptococcus pneumoniae (PRSP) and vancomycin-resistant enterococci (VRE) have been a serious problem in the medical community. Recently, the new quinolone antibacterials are highly successful class of antibacterial therapeutic field, however, the increased isolation of quinolone-resistant bacteria above them has become a normal outcome. These problems of multi-drug resistance have been the driving force for the development of newer quinolones. The next gereration of quinolone antibacterial agents will be potent against multi-drug resistant bacteria, such as MRSA, and provide a lower rate of emergence in resistance. Further, they should have favorable safety profiles to reduce the adverse reactions. The future of quinolones as the ultimate in pharmaceuticals must be handled cautiously if they are to realize their potential in the medical community.

Quinolone-DNA Interaction: Sequence-Dependent Binding to Single-Stranded DNA Reflects the Interaction within the Gyrase-DNA Complex

Abstract: We have investigated the interaction of quinolones with DNA by a number of methods to establish whether a particular binding mode correlates with quinolone potency. The specificities of the quinolone-mediated DNA cleavage reaction of DNA gyrase were compared for a number of quinolones. Two patterns that depended on the potency of the quinolone were identified. Binding to plasmid DNA was examined by measuring the unwinding of pBR322 by quinolones; no correlation with quinolone potency was observed. Quinolone binding to short DNA oligonucleotides was measured by surface plasmon resonance. The quinolones bound to both single- and double-stranded oligonucleotides in an Mg 2+ -dependent manner. Quinolones bound to single-stranded DNA with a higher affinity, and the binding exhibited sequence dependence; binding to double-stranded DNA was sequence independent. The variations in binding in the presence of metal ions showed that Mg 2+ promoted tighter, more specific binding to single-stranded DNA than softer metal ions (Mn 2+ and Cd 2+ ). Single-stranded DNA binding by quinolones correlated with the in vitro quinolone potency, indicating that this mode of interaction may reflect the interaction of the quinolone with DNA in the context of the gyrase-DNA complex.

Chromosomally encoded gyrase inhibitor GyrI protects Escherichia coli against DNA-damaging agents

Abstract: DNA gyrase, a type II topoisomerase, is the sole supercoiling activity in the cell and is essential for cell survival. There are two proteinaceous inhibitors of DNA gyrase that are plasmid-borne and ensure maintenance of the plasmids in bacterial populations. However, the physiological role of GyrI, an inhibitor of DNA gyrase encoded by the Escherichia coli genome, has been elusive. Previously, we have shown that GyrI imparts resistance against microcin B17 and CcdB. Here, we find that GyrI provided partial/limited protection against the quinolone class of gyrase inhibitors but had no effect on inhibitors that interfere with the ATPase activity of the enzyme. Moreover, GyrI negated the effect of alkylating agents, such as mitomycin C and N-methyl-N-nitro-N-nitrosoguanidine, that act independently of DNA gyrase. Hence, in vivo, GyrI appears to be involved in reducing DNA damage from many sources. In contrast, GyrI is not effective against lesions induced by ultraviolet radiation. Furthermore, the expression of GyrI does not significantly alter the topology of DNA. Thus, although isolated as an inhibitor of DNA gyrase, GyrI seems to have a broader role in vivo than previously envisaged.

Topoisomerase IV mutations in quinolone-resistant salmonellae selected in vitro.

Abstract: The development of high-level fluoroquinolone resistance has rarely been observed in salmonellae and, in contrast to other Gram-negative bacteria mutations affecting topoisomerase IV, a known secondary target of quinolones in Escherichia coli has not been described except for one recent report. The present study used quinolone-susceptible field isolates representing epidemiologically relevant serovars and phage types Salmonella Hadar and Salmonella Typhimurium DT104 and DT204c to select fluoroquinolone-resistant mutants in vitro. Three selection steps were necessary to obtain high-level fluoroquinolone-resistant mutants (MICCip > or = 8 microg/ml). All first-step mutants examined had a single gyrA mutation (affecting either Ser83 or Asp87). Additional topoisomerase mutations affecting gyrA (Asp87), gyrB (Ser464), and parC (Gly78) were detected in second- and third-step mutants. Introducing into the respective mutants the corresponding plasmid-coded quinolone-susceptible allele of either gyrA, gyrB, or parC resulted in reduction of quinolone resistance, indicating a role for these mutations in quinolone resistance. In the presence of an inhibitor of RND-type efflux pumps, the susceptibilities to ciprofloxacin and chloramphenicol of second- and third-step mutants increased by two to four serial dilution steps, providing evidence that an efflux-mediated resistance mechanism contributes to the development of high-level fluoroquinolone resistance in salmonellae.

Incidence of quinolone resistance in strains of Salmonella isolated from poultry. cattle and pigs in Germany between 1998 and 2001

Abstract: This paper reports the susceptibility to the quinolone nalidixic acid and the fluoroquinolone ciprofloxacin of 14,514 strains of Salmonella isolated in Germany from poultry, cattle and pigs between 1998 and 2001. Quinolone-resistant salmonellae were most frequently isolated from poultry, with a prevalence of 10.2 to 16-8 per cent. Poultry-associated serotypes, such as Salmonella Paratyphi B (d-tartrate positive), Salmonella Hadar and Salmonella Virchow, had the highest prevalence of quinolone resistance, ranging between 35 and 74 per cent. All the nalidixic acid-resistant strains also had a reduced susceptibility to ciprofloxacin, with minimum inhibitory concentrations (MICS) of 0.125 to 2 μg/ml. A comparison of the Mics for ciprofloxacin of the strains of these poultry-associated serotypes and Salmonella Enteritidis phage type 4 isolated in 1998/99 and 2000/01 indicated that there had been a shift towards higher Mic values of up to 2 μg/mI. The quinolone resistance-determining region (QRDR) of the gyrA gene and the homologue region of the parC gene of 31 selected strains were sequenced. Several different amino acid changes were observed in gyrA of the quinolone-resistant isolates at positions 83 and 87, but no substitutions were observed in parC .

Topoisomerase Targeting with and Resistance to Gemifloxacin in Staphylococcus aureus

Abstract: Gemifloxacin, a novel quinolone with potent activity against Staphylococcus aureus, was 8- to 16-fold more active against wild-type S. aureus than ciprofloxacin. The two- to fourfold increase in the MIC of gemifloxacin in genetically defined grlBA mutants and the twofold increase in a single gyrA mutant, supported by the low frequency of selection of resistant mutants at twice the MIC (7.4 x 10(-11) to 1.1 x 10(-10)), suggested similar targeting of the two enzymes by gemifloxacin. Dual mutations in both gyrase and topoisomerase IV caused a 64- to 128-fold increase in the MIC of gemifloxacin, similar to that seen with ciprofloxacin. Gemifloxacin also had similar activity in vitro against topoisomerase IV and gyrase purified from S. aureus (50% inhibitory concentrations of 0.25 and 0.31 micro g/ml, respectively). This activity was 10- to 20-fold higher than that of ciprofloxacin for topoisomerase IV and 33-fold higher than that for gyrase. In contrast to the in vitro findings, only topoisomerase IV mutants were selected in first-step mutants. Overexpression of the NorA efflux pump had a minimal effect on resistance to gemifloxacin, and a mutation in the promoter region of the gene for NorA was selected only in the sixth step of serial selection of mutants. Our data show that although gemifloxacin targets purified topoisomerase IV and gyrase similarly in vitro, topoisomerase IV is the preferred target in the bacteria. Selection of novel resistance mutations in grlA requires further expansion of quinolone-resistance-determining regions, and their study may provide increased insight into enzyme-quinolone interactions.

Discovery of (3S)-amino-(4R)-ethylpiperidinyl quinolones as potent antibacterial agents with a broad spectrum of activity and activity against resistant pathogens.

Abstract: Novel quinolone antibacterial agents bearing (3S)-amino-(4R)-ethylpiperidines were designed by using low energy conformation analysis and synthesized by applying a conventional coupling reaction of the quinolone nuclei with new piperidine side chains. These compounds were tested in MIC assays and found to be highly potent against Gram-positive and Gram-negative organisms. In particular, the new compounds exhibited high activity against the resistant pathogens Staphylococcus aureus (MRCR) and Streptococcus pneumoniae (PR). Importantly, when the (3S)-amino-(4R)-ethylpiperidinyl quinolones were compared with marketed quinolones sharing the same quinolone nuclei but different side chains at the C-7 position, the new quinolones showed superior activity against Gram-positive organisms, including resistant pathogens.

The history of quinolones

Abstract: The first quinolone emerged in the early 1960s, with the isolation of 7-chloro-l-ethyl-1, 4-dihydro-4-oxoquinoline-3-carboxylic acid, a by-product of the commercial preparation of chloroquine This compound was found to have anti-bacterial activity and was subsequently modified to produce nalidixic acid, a 1, 8-naphthyridine [1] (see Fig 1)

ATP-Bound Conformation of Topoisomerase IV: a Possible Target for Quinolones in Streptococcus pneumoniae

Abstract: Topoisomerase IV, a C2E2 tetramer, is involved in the topological changes of DNA during replication. This enzyme is the target of antibacterial compounds, such as the coumarins, which target the ATP binding site in the ParE subunit, and the quinolones, which bind, outside the active site, to the quinolone resistance-determining region (QRDR). After site-directed and random mutagenesis, we found some mutations in the ATP binding site of ParE near the dimeric interface and outside the QRDR that conferred quinolone resistance to Streptococcus pneumoniae, a bacterial pathogen. Modeling of the N-terminal, 43-kDa ParE domain of S. pneumoniae revealed that the most frequent mutations affected conserved residues, among them His43 and His103, which are involved in the hydrogen bond network supporting ATP hydrolysis, and Met31, at the dimeric interface. All mutants showed a particular phenotype of resistance to fluoroquinolones and an increase in susceptibility to novobiocin. All mutations in ParE resulted in resistance only when associated with a mutation in the QRDR of the GyrA subunit. Our models of the closed and open conformations of the active site indicate that quinolones preferentially target topoisomerase IV of S. pneumoniae in its ATP-bound closed conformation.

Unique Biological Properties and Molecular Mechanism of 5,6-Bridged Quinolones

Abstract: We have characterized an early series of 5,6-bridged dioxinoquinolones which behaved strikingly different from typical quinolones. The 5,6-bridged dioxinoquinolones inhibited Escherichia coli DNA gyrase supercoiling activity but, unlike typical quinolones, failed to stimulate gyrase-dependent cleavable complex formation. Analogous unsubstituted compounds stimulated cleavable complex formation but were considerably less potent than the corresponding 5,6-bridged compounds. Consistent with a previous report (M. Antoine et al., Chim. Ther. 7:434-443, 1972) and contrary to established quinolone SAR trends, a compound with an N-1 methyl substitution (PGE-8367769) was more potent than its analog with an N-1 ethyl substitution (PGE-6596491). PGE-8367769 was shown to antagonize ciprofloxacin-mediated cleavable complex formation in a dose-dependent manner, suggesting an interaction with the gyrase-DNA complex that overlaps that of ciprofloxacin. Resistance to PGE-8367769 in E. coli was found to arise through missense mutations in gyrA, implicating DNA gyrase as the primary antibacterial target. Notably, only 1 of 15 distinct mutations selected on PGE-8367769 (D87G) has previously been implicated in quinolone resistance in E. coli. The remaining 14 mutations (E16V, G31V, R38L, G40A, Y50D, V70A, A84V, I89L, M135T, G173S, T180I, F217C, P218T, and F513C) have not been previously reported, and most were located outside of the traditional quinolone resistance-determining region. These novel GyrA mutations decreased sensitivity to 5,6-bridged dioxinoquinolones by four- to eightfold, whereas they did not confer resistance to other quinolones such as ciprofloxacin, clinafloxacin, or nalidixic acid. These results demonstrate that the 5,6-bridged quinolones act via a mechanism that is related to but qualitatively different from that of typical quinolones.

Pharmacodynamics of Quinolone Antimicrobial Agents

Abstract: This chapter focuses on the pharmacodynamic characteristics of the quinolone antimicrobials in in vitro models, in animal infection models, and in humans. It demonstrates that there are many more similarities in results among the various models than there are differences. Numerous in vitro and in vivo studies have demonstrated that the quinolone antimicrobial agents exhibit concentration-dependent killing across a wide range of concentrations. A variety of different parameters have been used to characterize the killing characteristics of the quinolone antimicrobials. There are a variety of in vitro and in vivo persistent effects that have been characterized for the quinolone antimicrobial agents. The first study correlating pharmacokinetic (PK)/pharmacodynamic (PD) parameters of the fluoroquinolones with clinical response in humans was published by Peloquin. In this study, which evaluated intravenous ciprofloxacin in seriously ill patients with lower respiratory tract infections, time above MIC was reported to be the important parameter for eradication of the organism from respiratory secretions. Knowledge of the major PK/PD parameter determining efficacy and the magnitude of that target required for efficacy of specific pathogens has proven to be helpful for developing new quinolone agents, predicting the activity of new quinolone formulations, developing guideline recommendations, and establishing susceptibility and resistance breakpoints for susceptibility testing. Initial studies suggest that pharmacodynamics can also be important in preventing the emergence of resistance. More in vitro, animal and human studies are necessary to fully document the usefulness of pharmacodynamic evaluation in optimizing therapy with the quinolone antimicrobial agents.